AGRIN-INDUCED ACETYLCHOLINE-RECEPTOR CLUSTERING IN MAMMALIAN MUSCLE REQUIRES TYROSINE PHOSPHORYLATION

Agrin is thought to be the nerve-derived factor that initiates acetylc holine receptor (AChR) clustering at the developing neuromuscular junc tion. We have investigated the signaling pathway in mouse C2 myotubes and report that agrin induces a rapid but transient tyrosine phosphory lation of the AChR beta subunit. As the beta-subunit tyrosine phosphor ylation occurs before the formation of AChR clusters, it may serve as a precursor step in the clustering mechanism. Consistent with this, we observed that tyrosine phosphorylation of the beta subunit correlated precisely with the presence or absence of clustering under several ex perimental conditions. Moreover, two tyrosine kinase inhibitors, herbi mycin and staurosporine, that blocked beta-subunit phosphorylation als o blocked agrin-induced clustering. Surprisingly, the inhibitors also dispersed preformed AChR clusters, suggesting that the tyrosine phosph orylation of other proteins may be required for the maintenance of rec eptor clusters. These findings indicate that in mammalian muscle, agri n-induced AChR clustering occurs through a mechanism that requires tyr osine phosphorylation and may involve tyrosine phosphorylation of the AChR itself.